Analyzing clinical samples such as serum and urine with ISE devices involves multiple readings of a test solution using a plurality of ion specific analyte electrodes. Generally, an analyte electrode responds to a particular ion activity. In some ISE systems, generally termed indirect ISE systems, the test solution is mixed with a diluent producing an assay mixture. Use of diluted test solutions reduces the amount of test solution required and reduces the detrimental effect of materials such as protein that may be present at high concentration in some test solutions. In some systems, different aliquots of the same test solution or assay mixture may be applied to individual devices having different specific electrodes. Other systems combine several electrodes in one measuring device determining multiple ion activities from a single aliquot of test solution. Existing multi-electrode ISE devices, however, may use bulky configurations to provide the appropriate fluid flow over a plurality of electrodes and may require relatively large amounts of test solution to perform accurate mixing and measurements.
Generally, an ion selective electrode (ISE) is a sensor that responds to the activity of a specific ion in a solution by electrical means. The sensor, in cooperation with other components, translates the activity of a specific ion into a measurable electrical potential. The sensing portion of the electrode is usually made of an ion-specific membrane. A reference electrode completes an electric circuit for measuring the electrical potential. Commonly, a grounding connection ties the solution to system electrical ground to minimize electrical noise and to bring the potential into a range compatible with measurement electronics. The voltage developed between the analyte electrode and the reference electrode may be used to determine the activity of ions in the solution.
Ion-selective electrodes are widely used in biochemical and biophysical assays for measurements of ionic concentration in aqueous solutions. Clinical electrolyte assays frequently employ ISEs to determine multiple ion activities in serum, plasma, cerebrospinal fluid, or urine. Such test solutions are usually available in very limited quantities. Existing ISE systems use as small a volume of such test solutions as practical, but the volume used is still quite large; some systems use 65 microliters or more. This is ten to twenty times the volume used for other routine clinical assays.
Generally, a clinical ISE system constantly interchanges the fluids that fill it, presenting to the electrodes in turn a first assay mixture, a reference solution, a second assay mixture, a reference solution, etc. Any amount of a previous fluid that remains within the system must be a small enough proportion of the volume of the next fluid that it contributes no more than a negligible effect on the measured result. Thus the volume of an ISE system has a two-fold influence on the amount of test solution required; first, enough assay mixture must be available to fill the volume; second, the volume of assay mixture must be much greater than any residual volume of a prior fluid. The amount of residual fluid depends upon the area of wetted surface and upon the geometry of that surface. Tight corners, complex shapes exposed to wetting, unswept dead volumes, and rough surfaces all increase the amount of residual fluid and require more assay mixture, and hence more of the scarce test solution.
One test solution reduction strategy commonly employed is to expose a single aliquot of test solution to multiple electrodes. Several drawbacks may exist in existing multi-electrode ISE testing devices. For example, some existing systems may use separate, disjoint components to form the multi-electrode ISE assay system. Generally, fluid connectors and tubing connect disjoint elements of existing ISE systems to interconnect a working system. These disjoint systems may require large amounts of test solution to fill the fluid connectors and tubing between the disjoint elements. Still more test solution may be required to rinse prior fluids from the tubing and from dead spaces in fluid connectors.
Moreover, some existing systems are inefficient in preparing a sample for testing. In particular, mixing precision and efficiency may be problematic in some indirect ISE systems. Some ISE systems may use a mechanical stir bar to mix a test solution with a diluent. Stir bars and similar mixers with their associated drive mechanisms add mechanical and operational costs to an ISE system. Stir bars also need thorough cleaning between applications of different test solutions to prevent cross-contamination (i.e. carryover) that may skew test results. Further, the stir bar requires a minimum amount of fluid to mix effectively, thereby placing additional volume constraints on scarce test samples.
Other clinical ISE systems use a closed injection port where a stream of diluent interacts with a stream of test solution delivered by a probe sealed into the injection port. Fluid mixing in such systems is dependent on the interpenetration of liquid streams from two sources, a process directly opposed by fluid viscosity. Relatively high fluid injection velocities are required to produce this mixing. Maintaining the desired ratio of test solution to diluent under such conditions generally requires a pump that is inherently ratiometric, such as a multi-chamber positive displacement ratio pump. Such pumps have proven problematic, in part due to leakage at multiple seals that produce undesired fluid junctions that upset electrode potentials. All surfaces of the closed injection port need thorough cleaning between applications of different test solutions to prevent carryover that may skew test results. Further, the probe seal in such systems is a wear part that may require frequent maintenance.
Another problem with some existing systems may involve the grounding configuration of these systems. As described above, typical ISE systems use a ground connection to bring the assay mixture potential into measurable range for the electronics. Existing ISE systems may use a solution ground that is a conductive tube suspended and immersed in the assay mixture or a grounding plate disposed in flow channels between the analyte electrodes. The ground may be anchored by a coupling that requires glue. The glue may contaminate the assay mixture, causing unwanted electrical noise, drift, or skewing in the measurements of the ISE system. Grounding plates suspended in flow channels may create dead volumes and crevices that are difficult to clean, increasing the volume of test solutions needed. Further, the solution ground may be difficult to insert, assemble, or connect, adding to the expense of construction of an ISE system.
Yet another issue with some existing systems is the requirement of a ratio pump or other multi-chamber pump to precisely control the movement of test solution, diluent, reference buffer, and other solutions through the flow cell from a dilution cup. Due to the pump design, the solutions are able to wick from one chamber to another, thereby contaminating the solutions in the chambers. Because not all solutions are used at the same time, the solutions are repeatedly cycled (e.g. diluent is aspirated and dispensed back to its source) and therefore more likely to contaminate the entire system. The wicking between chambers may also produce a half-cell; the resulting potential may skew the ISE measurements. Ratio pumps and other multi-chamber pumps may be expensive and difficult to maintain. Because of the number of components of these pumps and complexity of operation, mechanical failures may occur frequently. These failures may also be difficult and time consuming to repair.